1. Biochemistry and Chemical Biology
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RNA-guided retargeting of Sleeping Beauty transposition in human cells

  1. Adrian Kovač
  2. Csaba Miskey
  3. Michael Menzel
  4. Esther Grueso
  5. Andreas Gogol-Döring
  6. Zoltan Ivics  Is a corresponding author
  1. Paul Ehrlich Institute, Germany
  2. University of Applied Sciences, Germany
Research Article
  • Cited 7
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Cite this article as: eLife 2020;9:e53868 doi: 10.7554/eLife.53868

Abstract

An ideal tool for gene therapy would enable efficient gene integration at predetermined sites in the human genome. Here we demonstrate biased genome-wide integration of the Sleeping Beauty (SB) transposon by combining it with components of the CRISPR/Cas9 system. We provide proof-of-concept that it is possible to influence the target site selection of SB by fusing it to a catalytically inactive Cas9 (dCas9) and by providing a single guide RNA (sgRNA) against the human Alu retrotransposon. Enrichment of transposon integrations was dependent on the sgRNA, and occurred in an asymmetric pattern with a bias towards sites in a relatively narrow, 300-bp window downstream of the sgRNA targets. Our data indicate that the targeting mechanism specified by CRISPR/Cas9 forces integration into genomic regions that are otherwise poor targets for SB transposition. Future modifications of this technology may allow the development of methods for specific gene insertion for precision genetic engineering.

Data availability

DNA sequence data generated and analysed during this study are included in the manuscript and Source Data Files 1-4.

Article and author information

Author details

  1. Adrian Kovač

    Medical Biotechnology, Paul Ehrlich Institute, Langen, Germany
    Competing interests
    No competing interests declared.
  2. Csaba Miskey

    Medical Biotechnology, Paul Ehrlich Institute, Langen, Germany
    Competing interests
    No competing interests declared.
  3. Michael Menzel

    Bioinformatics, University of Applied Sciences, Giessen, Germany
    Competing interests
    No competing interests declared.
  4. Esther Grueso

    Medical Biotechnology, Paul Ehrlich Institute, Langen, Germany
    Competing interests
    No competing interests declared.
  5. Andreas Gogol-Döring

    Bioinformatics, University of Applied Sciences, Giessen, Germany
    Competing interests
    No competing interests declared.
  6. Zoltan Ivics

    Medical Biotechnology, Paul Ehrlich Institute, Langen, Germany
    For correspondence
    zoltan.ivics@pei.de
    Competing interests
    Zoltan Ivics, Patent applications around targeted transposon integration technology(Patent Nos. EP1594971B1, EP1594972B1 and EP1594973B1)..
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-7803-6658

Funding

No external funding was received for this work.

Reviewing Editor

  1. Jeffrey Essner, Iowa State University, United States

Publication history

  1. Received: November 22, 2019
  2. Accepted: March 5, 2020
  3. Accepted Manuscript published: March 6, 2020 (version 1)
  4. Version of Record published: March 17, 2020 (version 2)

Copyright

© 2020, Kovač et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

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